Publications by authors named "Dan W Nowakowski"

7 Publications

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A Combinatorial MAP Code Dictates Polarized Microtubule Transport.

Dev Cell 2020 04 27;53(1):60-72.e4. Epub 2020 Feb 27.

Department of Molecular and Cellular Biology, University of California, Davis, Davis, CA 95616, USA. Electronic address:

Many eukaryotic cells distribute their intracellular components asymmetrically through regulated active transport driven by molecular motors along microtubule tracks. While intrinsic and extrinsic regulation of motor activity exists, what governs the overall distribution of activated motor-cargo complexes within cells remains unclear. Here, we utilize in vitro reconstitution of purified motor proteins and non-enzymatic microtubule-associated proteins (MAPs) to demonstrate that MAPs exhibit distinct influences on the motility of the three main classes of transport motors: kinesin-1, kinesin-3, and cytoplasmic dynein. Further, we dissect how combinations of MAPs affect motors and unveil MAP9 as a positive modulator of kinesin-3 motility. From these data, we propose a general "MAP code" that has the capacity to strongly bias directed movement along microtubules and helps elucidate the intricate intracellular sorting observed in highly polarized cells such as neurons.
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http://dx.doi.org/10.1016/j.devcel.2020.01.029DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC7181406PMC
April 2020

Microtubules gate tau condensation to spatially regulate microtubule functions.

Nat Cell Biol 2019 09 2;21(9):1078-1085. Epub 2019 Sep 2.

Department of Molecular and Cellular Biology, University of California, Davis, Davis, CA, USA.

Tau is an abundant microtubule-associated protein in neurons. Tau aggregation into insoluble fibrils is a hallmark of Alzheimer's disease and other types of dementia, yet the physiological state of tau molecules within cells remains unclear. Using single-molecule imaging, we directly observe that the microtubule lattice regulates reversible tau self-association, leading to localized, dynamic condensation of tau molecules on the microtubule surface. Tau condensates form selectively permissible barriers, spatially regulating the activity of microtubule-severing enzymes and the movement of molecular motors through their boundaries. We propose that reversible self-association of tau molecules, gated by the microtubule lattice, is an important mechanism of the biological functions of tau, and that oligomerization of tau is a common property shared between the physiological and disease-associated forms of the molecule.
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http://dx.doi.org/10.1038/s41556-019-0375-5DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC6748660PMC
September 2019

De novo variant in KIF26B is associated with pontocerebellar hypoplasia with infantile spinal muscular atrophy.

Am J Med Genet A 2018 12 27;176(12):2623-2629. Epub 2018 Aug 27.

Division of Newborn Medicine, Boston Children's Hospital, Harvard Medical School, Boston, Massachusetts.

KIF26B is a member of the kinesin superfamily with evolutionarily conserved functions in controlling aspects of embryogenesis, including the development of the nervous system, though its function is incompletely understood. We describe an infant with progressive microcephaly, pontocerebellar hypoplasia, and arthrogryposis secondary to the involvement of anterior horn cells and ventral (motor) nerves. We performed whole exome sequencing on the trio and identified a de novo KIF26B missense variant, p.Gly546Ser, in the proband. This variant alters a highly conserved amino acid residue that is part of the phosphate-binding loop motif and motor-like domain and is deemed pathogenic by several in silico methods. Functional analysis of the variant protein in cultured cells revealed a reduction in the KIF26B protein's ability to promote cell adhesion, a defect that potentially contributes to its pathogenicity. Overall, KIF26B may play a critical role in the brain development and, when mutated, cause pontocerebellar hypoplasia with arthrogryposis.
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http://dx.doi.org/10.1002/ajmg.a.40493DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC6481602PMC
December 2018

NGP 555, a γ-Secretase Modulator, Lowers the Amyloid Biomarker, Aβ in Cerebrospinal Fluid while Preventing Alzheimer's Disease Cognitive Decline in Rodents.

Alzheimers Dement (N Y) 2017 Jan;3(1):65-73

NeuroGenetic Pharmaceuticals, Inc., Del Mar, CA 92014.

Introduction: Alzheimer's disease (AD) is defined by the progressive accumulation of amyloid plaques and neurofibrillary tangles in the brain which precedes cognitive decline by years.

Methods: Using amyloid biomarkers, chemical modeling, mouse behavioral models, and drug development techniques we investigate the properties of NGP 555, a clinical-stage γ-secretase modulator.

Results: NGP 555 shifts amyloid peptide production to the smaller, non-aggregating forms of amyloid. Our preclinical studies show beneficial effects on amyloid biomarkers, pathology, and cognition. NGP 555 has successfully completed chemistry, pharmacology, toxicity, metabolism, and safety studies.

Discussion: Abundant data support Aβ as a target for prophylactic or early-stage intervention therapies in AD. The γ-secretase modulator, NGP 555 is being actively developed in human clinical trials for the prevention of Alzheimer's disease with the overall aim to achieve an appropriate balance of potency/efficacy on reducing the toxic forms of amyloid versus safety.
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http://dx.doi.org/10.1016/j.trci.2016.09.003DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC5421551PMC
January 2017

APC is an RNA-binding protein, and its interactome provides a link to neural development and microtubule assembly.

Cell 2014 Jul;158(2):368-382

Department of Cell Biology and Program in Neuroscience, Harvard Medical School, Boston, MA 02115, USA. Electronic address:

Adenomatous polyposis coli (APC) is a microtubule plus-end scaffolding protein important in biology and disease. APC is implicated in RNA localization, although the mechanisms and functional significance remain unclear. We show APC is an RNA-binding protein and identify an RNA interactome by HITS-CLIP. Targets were highly enriched for APC-related functions, including microtubule organization, cell motility, cancer, and neurologic disease. Among the targets is β2B-tubulin, known to be required in human neuron and axon migration. We show β2B-tubulin is synthesized in axons and localizes preferentially to dynamic microtubules in the growth cone periphery. APC binds the β2B-tubulin 3' UTR; experiments interfering with this interaction reduced β2B-tubulin mRNA axonal localization and expression, depleted dynamic microtubules and the growth cone periphery, and impaired neuron migration. These results identify APC as a platform binding functionally related protein and RNA networks, and suggest a self-organizing model for the microtubule to localize synthesis of its own subunits.
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http://dx.doi.org/10.1016/j.cell.2014.05.042DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC4133101PMC
July 2014

Type III neuregulin 1 regulates pathfinding of sensory axons in the developing spinal cord and periphery.

Development 2011 Nov;138(22):4887-98

Department of Cell Biology, Harvard Medical School, Boston, MA 02115, USA.

Sensory axons must develop appropriate connections with both central and peripheral targets. Whereas the peripheral cues have provided a classic model for neuron survival and guidance, less is known about the central cues or the coordination of central and peripheral connectivity. Here we find that type III Nrg1, in addition to its known effect on neuron survival, regulates axon pathfinding. In type III Nrg1(-/-) mice, death of TrkA(+) nociceptive/thermoreceptive neurons was increased, and could be rescued by Bax elimination. In the Bax and type III Nrg1 double mutants, axon pathfinding abnormalities were seen for TrkA(+) neurons both in cutaneous peripheral targets and in spinal cord central targets. Axon guidance phenotypes in the spinal cord included penetration of axons into ventral regions from which they would normally be repelled by Sema3A. Accordingly, sensory neurons from type III Nrg1(-/-) mice were unresponsive to the repellent effects of Sema3A in vitro, which might account, at least in part, for the central projection phenotype, and demonstrates an effect of type III Nrg1 on guidance cue responsiveness in neurons. Moreover, stimulation of type III Nrg1 back-signaling in cultured sensory neurons was found to regulate axonal levels of the Sema3A receptor neuropilin 1. These results reveal a molecular mechanism whereby type III Nrg1 signaling can regulate the responsiveness of neurons to a guidance cue, and show that type III Nrg1 is required for normal sensory neuron survival and axon pathfinding in both central and peripheral targets.
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http://dx.doi.org/10.1242/dev.072306DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC3201659PMC
November 2011

A protein complex containing Mdm10p, Mdm12p, and Mmm1p links mitochondrial membranes and DNA to the cytoskeleton-based segregation machinery.

Mol Biol Cell 2003 Nov 17;14(11):4618-27. Epub 2003 Sep 17.

Department of Anatomy and Cell Biology, Columbia University College of Physicians and Surgeons, New York, New York 10032, USA.

Previous studies indicate that two proteins, Mmm1p and Mdm10p, are required to link mitochondria to the actin cytoskeleton of yeast and for actin-based control of mitochondrial movement, inheritance and morphology. Both proteins are integral mitochondrial outer membrane proteins. Mmm1p localizes to punctate structures in close proximity to mitochondrial DNA (mtDNA) nucleoids. We found that Mmm1p and Mdm10p exist in a complex with Mdm12p, another integral mitochondrial outer membrane protein required for mitochondrial morphology and inheritance. This interpretation is based on observations that 1) Mdm10p and Mdm12p showed the same localization as Mmm1p; 2) Mdm12p, like Mdm10p and Mmm1p, was required for mitochondrial motility; and 3) all three proteins coimmunoprecipitated with each other. Moreover, Mdm10p localized to mitochondria in the absence of the other subunits. In contrast, deletion of MMM1 resulted in mislocalization of Mdm12p, and deletion of MDM12 caused mislocalization of Mmm1p. Finally, we observed a reciprocal relationship between the Mdm10p/Mdm12p/Mmm1p complex and mtDNA. Deletion of any one of the subunits resulted in loss of mtDNA or defects in mtDNA nucleoid maintenance. Conversely, deletion of mtDNA affected mitochondrial motility: mitochondria in cells without mtDNA move 2-3 times faster than mitochondria in cells with mtDNA. These observations support a model in which the Mdm10p/Mdm12p/Mmm1p complex links the minimum heritable unit of mitochondria (mtDNA and mitochondrial outer and inner membranes) to the cytoskeletal system that drives transfer of that unit from mother to daughter cells.
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http://dx.doi.org/10.1091/mbc.e03-04-0225DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC266777PMC
November 2003